SUMMARY and ABSTRACT Glia comprise approximately 60% of the cellular constituency of central nervous system (CNS), playing diverse roles in the functioning CNS and a host of neurological disorders. Development of glial cell lineages proceeds along a tightly regulated program that involves patterning, generation of diverse cells, differentiation, and myelination. This cascade of developmental events is particularly vulnerable to neonatal hypoxic brain injury, which leads to profound loss of myelinating oligodendrocytes (OLs), extensive white matter damage, culminating in neuronal dysfunction. Despite the robust regenerative capacity of OLs, the underlying mechanisms mediating hypomyelination and the subsequent defects in neural circuits after hypoxic injury remain poorly defined. Moreover, myelination continues throughout early adulthood, which also renders the CNS susceptible to insults causing late-onset neurodegeneration. Therefore, the overarching goal of this application is to define new genes and pathways that drive OL maturation during development and repair, and pinpoint potential targetable pathways for white matter disorders. Previously, we identified Daam2 (Disheveled associated activator of morphogenesis 2) as a pivotal regulator of OL myelination and repair, and recently discovered that Daam2 governs OL differentiation through ubiquitination of the hypoxia regulator VHL (von Hippel-Lindau). Moreover, we discovered that Daam2 is regulated by two E3 ligases, Nedd4 (Neural precursor cell expressed developmentally down-regulated protein 4) and Trim9 (Tripartite Motif Containing 9), which in turn govern VHL ubiquitination and OL differentiation. These observations raise two key questions that we will pursue in this proposal: 1) how does Daam2 modulate VHL-HIF signaling in OLs? and 2) how is the ubiquitin-mediated Daam2 degradation controlled in OLs? By understanding the in-depth mechanisms by which Daam2 operates, we will establish Daam2 inhibition as a clinically significant and actionable strategy for the treatment of white matter injury. To answer these key questions, we will first define the reciprocal relationship between Daam2 and VHL during OL development and white matter injury (Aim 1). These studies will define the Daam2-VHL axis as a pivotal regulator of OL development, while revealing novel connections between Wnt signaling and hypoxic pathway during OL myelination. Next, we will determine Daam2 proteasomal degradation pathways in OLs (Aim 2). Upon completion, these studies will define how Daam2 is regulated by target E3 ligases, and identify Nedd4 and Trim9 as novel regulators of OL myelination. A mechanistic understanding of Daam2-VHL axis function in oligodendrocyte repair after injury will shed light on cellular vulnerability to white matter injury and ultimately point to new venues for therapeutic development to stimulate OL remyelination.